Objective To evaluate the in vitro anti-hypertrophic effect of total Glycosides of Ranunculus Japonius (TGRJ).
Methods Neonatal rat cardiomyocytes were cultured and hypertrophy was induced by adminis- trating isoproterenol (ISO, 10 μmol/L) or angiotensin Ⅱ (Ang Ⅱ, 1 μmol/L) for 48 hours. In the treatment groups, cells were pretreated with TGRJ (0.3 g/L) for 30 minutes prior to hypertrophic stimuli. The anti-hypertrophic effects of TGRJ were examined by measuring cell size, total protein content, and protein synthesis. Intracellular free Ca²⁺ concentration ([Ca²⁺]_i) was evaluated using fluorescence dye Fura-2/AM. Sacroplasmic/endoplasmic reticulum Ca²⁺ ATPase 2a (SERCA2a), atrial natriuretic peptide (ANP), B-type natriuretic peptide (BNP), and beta-myosin heavy chain (β-MHC) protein expression levels were measured by Western blotting . SERCA2a activity was assayed by p-nitrophenal phosphate disodium salt hexahydrate method.
Results Increased cell size, total protein content, and protein synthesis following ISO or Ang Ⅱ stimulation were significantly inhibited by pretreatment with TGRJ (all P<0.05). This anti-hypertrophic effect of TGRJ was confirmed by its suppressing effect on elevated expression of the three hypertrophic related genetic markers, ANP, BNP, and β-MHC. In addition, TGRJ inhibited ISO or Ang Ⅱ induced up-regulation of [Ca²⁺]_i under chronic but not acute conditions. And ISO or Ang Ⅱ induced down-regulation of SERCA2a expression and activity was also effectively rectified by TGRJ pretreatment.
Conclusions The results of present study suggested that TGRJ could prevent ISO or Ang Ⅱ induced cardiac hypertrophy through improving chronic [Ca²⁺]_i disorder, might via normalizing SERCA2a expression and activity.

Key words: total Glycosides of Ranunculus Japonius, cardiac hypertrophy, isoproterenol, angiotensin Ⅱ, Ca²⁺, sacroplasmic/endoplasmic reticulum Ca²⁺ ATPase 2a